Product details

Product details

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Purpose

Annexin V Apoptosis Detection Kit is based on the observation that soon after initiating apoptosis, cells translocate the membrane phosphatidyl-serine (PS) from the inner face of the plasma membrane to the cell surface. Once on the cell surface, PS can be easily detected by staining with a fluorescent conjugate of Annexin V, a protein that has a high affinity for PS. The one-step staining procedure takes only 10 minutes. Detection can be analyzed by flow cytometry or by fluorescence microscopy.

Sample Type

Biological Fluids

Detection Method

Fluorometric

Specificity

The assay is based on the observation that soon after initiating apoptosis, most cell types translocate the membrane phospholipid phosphatidylserine (PS) from the inner face of the plasma membrane to the cell surface. Once on the cell surface, PS can easily be detected by staining with a fluorescent conjugate of Annexin V, a protein that has a strong natural affinity for PS.

C. Detection by Fluorescence Microscopy 1. Place the cell suspension from Step A.5 on a glass slide. Cover the cells with a glass coverslip. For analyzing adherent cells, grow cells directly on a coverslip. Following incubation (A.5), invert coverslip on glass slide and visualize cells. The cells can also be washed and fixed in 2 % formaldehyde before visualization. (Cells must be incubated with Annexin V-Cy3 before fixation since any cell membrane disruption can cause nonspecific binding of Annexin V to PS on the inner surface of the cell membrane.) 2. Observe the cells under a fluorescence microscope using Cy5 filter, or FITC/Cy3/Cy5 triple band filter (Chroma Technology) if performing triple labeling using these dyes, or detect cells using a CCD camera. Cells that have bound Annexin V-Cy5 will show bright red-blue staining on the plasma membrane.

Segawa, Suzuki, Nagata: "Constitutive exposure of phosphatidylserine on viable cells." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 108, Issue 48, pp. 19246-51, 2011 (PubMed).

Breckenridge, Nguyen, Kuppig et al.: "The procaspase-8 isoform, procaspase-8L, recruited to the BAP31 complex at the endoplasmic reticulum." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 99, Issue 7, pp. 4331-6, 2002 (PubMed).